In Ireland, Italian ryegrass is on a small number of farms. However, growers who have Italian ryegrass have found this weed extremely difficult to control.
On farms with populations it should be treated as seriously as blackgrass, because both have much the same biology.
Italian ryegrass is a cross-pollinating species.It has an early autumn emergence pattern, between September and November. Plants can only emerge from seeds found within the top 5cm of soil. If left uncontrolled, they can produce up to 5,000 seeds per plant. Seeds have short dormancy (less than five years). Worryingly, ryegrass species will develop herbicide resistance faster than blackgrass. In Europe and in Britain, there are populations of Italian ryegrass resistant to ACCase, ALS, flufenacet, and glyphosate herbicides.
It is important that any plants not controlled each year do not incur a high seed return, which would drive rapid resistance development. Cases of resistance in Italian ryegrass have already been detected in Ireland.
In this article, I report the first cases of herbicide-resistant Italian ryegrass from Ireland, the type of resistance, which is evolving, and the integrated weed management (IWM) strategies needed to eliminate or control these resistant strains.
These measures will help to contain the early problems and potentially prevent the expansion of its infestation in cereal fields.
Chemical options
Chemicals will be the first line of defence against an emerging problem but this needs a wide ranging and varied approach.
Glyphosate before sowing the crop to kill all emerged plants.Use pre-emergence or very early post-emergence herbicides like pendimethalin (Stomp Aqua)- or flufenacet (Firebird Met)-based products, prosulfocarb (Defy) or tri-allate (Avadex) in winter cereals. Use non-ACCase and non-ALS herbicide, eg Kerb in winter oilseed rape.Many post-emergence herbicides for spring wild oats are also very active on Italian ryegrass. •ACCase-Axial in barley or wheat (spring and winter).
• ACCase-Falcon, Stratos Ultra or Centurion Max in winter oilseed rape.
•ALS herbicides-Pacifica Plus, Broadway Star or Monolith in winter wheat.
Glyphosate use before harvesting to kill established plants but this will not destroy the seeds which have set.Confirmation of resistant
Italian ryegrass
Italian ryegrass populations from eight fields were collected prior to harvest in 2020 as a part of the ECT nationwide grass weed survey. Four of these populations (R1 to R4) from Tipperary, Cork and Meath, which appeared to exhibit resistance, were subjected to detailed dose-response studies where they were compared with a known sensitive population.
Plants were sprayed at the two- to three-leaf stage, with rates ranging from 0.25 to eight times the recommended field rates of ALS (Pacifica Plus and Broadway Star) and ACCase (Axial Pro, Falcon and Stratos Ultra) herbicides to quantify resistance levels (Figure 1).
Figure 1: Symptoms of susceptible and resistant (R1 to R4) populations of Italian ryegrass 28-days post the application of ALS (A and B) and ACCase (C to E) herbicides at dose rates from 0.25 to 8-times the recommended field rates (which are highlighted in red). The extent of the green vegetation in each row is indicative of the difficulty in killing that population with differing rates of individual herbicides.
The results in Figure 1 show:
Figures 1 A and B show that Pacifica Plus and Broadway Star were virtually ineffective in all populations, confirming that R1 to R4 were ALS-resistant.Figure 1 C shows that R2 and R3 survived Axial application, while R1 and R4 were totally killed by Axial at the lowest dose rate used.Figure 1 D shows that three populations (R1, R3 and R4) were totally killed by Falcon at the recommended field rate, but R2 was not adequately controlled.Figure 1 E shows that all four populations (R1 to R4) were totally killed by Stratos Ultra used at the recommended field rate. 
Figure 2: Pendimethalin-treated seedlings from a susceptible and two resistant populations (R1 and R4) had stunted shoots, indicating that they are highly pendimethalin-sensitive (ie no NTSR). In contrast, pendimethalin-treated plants from resistant populations R2 and R3 had developed normal shoots, indicating some degree of tolerance to pendimethalin and confirming potential NTSR.
In summary, populations R1 and R4 were ALS-resistant only. While populations R2 and R3 were ACCase- and ALS-resistant.
Nevertheless, ACCase-Stratos Ultra was highly effective on all four resistant populations. These findings demonstrate the difficulty in predicting cross-resistance, and stress the need to protect effective chemistry for critical use only.
Types of resistance evolving
in Irish Italian ryegrass
Following repeated application, Italian ryegrass develops herbicide resistance via two distinct mechanisms: (a) target-site resistance (TSR) where a plant changes the structure of its herbicide-binding site due to mutation(s) and this blocks herbicide activity or (b) non-target-site resistance (NTSR) where plants are able to degrade herbicides very rapidly before they reach the binding site.
To detect the involvement of TSR mutations, DNA was extracted from the resistant populations R1 to R4 using plant materials that survived ACCase/ALS herbicide application.
We found TSR as the key mechanism for ALS resistance in R1, R2 and R4. Even though R2 survived Axial or Falcon, no ACCase TSR detected. Similarly, R3 survived Pacifica Plus and Broadway Star, as well as Axial, but without ALS or ACCase TSR. These results speculate the possibility of NTSR in R2 and R3.To detect the involvement of NTSR or enhanced metabolism, we carried out a simple test using the pre-emergence herbicide pendimethalin, which is likely to affect the activity of other pre-emergence herbicides as well. However, resistance to pre-emergence herbicides used for grass weed control tends to be only partial and builds up relatively slowly.
Positive correlation between pendimethalin resistance and NTSR has been used for decades to screen UK grass weed populations.
A second test using a portable lateral flow device, which works on the same basis as a pregnancy test or COVID-19 antigen test, was used in parallel to detect the levels of herbicide-degrading enzymes GSTF1 (glutathione transferase phi (F class) 1) within 10 minutes (Figure 3).
Figure 3: Box plots of test line reading at 10 minutes from leaf protein extracted from Italian ryegrass plants. Each bar represents test line reading of five independent samples from each population. Different letter represents statistical significance (p<0.05). The test line reading of R3 was ˜2 times higher and statistically different to susceptible population, indicating NTSR development.
Higher levels of GSTF1 in the sample positively correspond to the existence of NTSR in the population.
Note: All NTSR work was performed at a leading UK institution, which has a unique molecular research capacity.
Using herbicide screening and molecular diagnostic tools, we identified and confirmed the development of NTSR alone or NTSR and TSR combinations in Irish Italian ryegrass populations (Table 1).
NTSR is a serious threat, as it affects multiple herbicides with different modes of action, leaving very limited chemical options for weed control.
High levels of control in these farms are needed to avoid further spread and buildup of resistance. All growers should address this weed using a zero-tolerance policy.
Resistance management and IWM
Remember, even though herbicides provide the most efficient and cost-effective means of managing weeds, resistance will make them obsolete if they are used as a sole weed control method without IWM.
Italian ryegrass is an increasing grass weed problem in Irish fields. It carries a very high risk of resistance development and some herbicide resistant strains have already been found in Irish fields.
Based on our knowledge of cross-resistance to key herbicides and the mechanisms involved, the IWM actions needed are as follows:
1 Farms where Italian ryegrass populations have confirmed or suspected herbicide resistance to ALS modes of action.
Pre-emergence herbicides in combination with cultural/non-chemical IWM like delayed autumn drilling, spring cropping, higher seed rates, ploughing, and hand-rogueing are critical to managing the soil seedbank population.No evidence of glyphosate resistance detected in R1 to R4. Therefore, glyphosate use on stubbles (at least 1.5 l/ha) may offer effective control but a follow-up treatment achieving high rates of control may be needed in the crop. For ALS-resistant populations (eg R1 and R4 in Table 1), adequate control may initially be achieved using ACCase (eg Axial Pro, Falcon and Stratos Ultra) herbicides.2 Farms where Italian ryegrass populations have confirmed or suspected herbicide resistance to ACCase and ALS modes of action.
Glyphosate use on stubbles (at least 1.5 l/ha) may offer effective control of young plants before planting.For ACCase/ALS-resistant populations (eg R2 and R3 in Table 1), ACCase-Stratos Ultra seems to be unaffected by NTSR and hence can be used to eliminate these multi-resistant types. However, the rotational restrictions around winter oilseed rape will limit the effectiveness of Stratos Ultra, hence the need to practice all and every cultural/non-chemical IWM to reduce the seed bank and prevent further seed return.General
prevention and weed management
In severely infested fields, extreme IWM measures such as cutting the crop for silage or spraying it off with glyphosate before seed return are important. Stale seedbed and cultivation: shallow cultivation immediately after harvest encourages germination of freshly shed seeds, which should be sprayed off with glyphosate prior to sowing. This technique is critical for direct drill or strip till, where seeds are retained on the soil surface. While ploughing can bury weed seeds to a depth from which they are unlikely to emerge, it can also bring previously buried seeds to the surface. Alternating the plough with a few years of minimum cultivation can reduce seedbank build-up.Delayed autumn drilling until mid-October will help to bypass the main autumn flush of Italian ryegrass and use higher seed rates to improve relative crop competitiveness.Spring cropping is a practical option where delayed autumn drilling is not possible or where weed pressure is significant
UK experience shows that hybrid barley can significantly reduce the level of Italian ryegrass seed return due to its higher competitive ability. Spring cropping is a practical option where delayed autumn drilling is not possible or where weed pressure is significant. However, a single spring crop may not be appropriate in fields with high soil seedbank populations. Including broadleaved crops in the rotation allows the use of other herbicide types (eg ACCase-Stratos Ultra). Hand-rogueing of small infestations or spraying off distinct patches prevent spreading within fields.Machine hygiene prevents weed seed movement within or between fields.Where infestations have occurred, pre-emergence herbicides should be followed by post-emergence herbicides applied to small actively growing plants. This practice should only be adopted in fields where weed pressure is low and ACCase/ALS resistance is not suspected.Finally, conduct resistance testing not only to determine whether the population is resistant, but also to identify the most effective chemistry that offers control for your populations.The presence of target site resistance and/or non-target site resistance has been confirmed in Irish Italian ryegrass populations.Increased vigilance, resistance testing and a zero-tolerance policy are critical to combat this weed.Do not use ACCase/ALS herbicides as a sole weed control method in consecutive crops due to its high resistance development risk.Use a wide range of cultural and non-chemical IWM options to protect effective herbicides for critical use and for the productivity of our cropping systems.
In Ireland, Italian ryegrass is on a small number of farms. However, growers who have Italian ryegrass have found this weed extremely difficult to control.
On farms with populations it should be treated as seriously as blackgrass, because both have much the same biology.
Italian ryegrass is a cross-pollinating species.It has an early autumn emergence pattern, between September and November. Plants can only emerge from seeds found within the top 5cm of soil. If left uncontrolled, they can produce up to 5,000 seeds per plant. Seeds have short dormancy (less than five years). Worryingly, ryegrass species will develop herbicide resistance faster than blackgrass. In Europe and in Britain, there are populations of Italian ryegrass resistant to ACCase, ALS, flufenacet, and glyphosate herbicides.
It is important that any plants not controlled each year do not incur a high seed return, which would drive rapid resistance development. Cases of resistance in Italian ryegrass have already been detected in Ireland.
In this article, I report the first cases of herbicide-resistant Italian ryegrass from Ireland, the type of resistance, which is evolving, and the integrated weed management (IWM) strategies needed to eliminate or control these resistant strains.
These measures will help to contain the early problems and potentially prevent the expansion of its infestation in cereal fields.
Chemical options
Chemicals will be the first line of defence against an emerging problem but this needs a wide ranging and varied approach.
Glyphosate before sowing the crop to kill all emerged plants.Use pre-emergence or very early post-emergence herbicides like pendimethalin (Stomp Aqua)- or flufenacet (Firebird Met)-based products, prosulfocarb (Defy) or tri-allate (Avadex) in winter cereals. Use non-ACCase and non-ALS herbicide, eg Kerb in winter oilseed rape.Many post-emergence herbicides for spring wild oats are also very active on Italian ryegrass. •ACCase-Axial in barley or wheat (spring and winter).
• ACCase-Falcon, Stratos Ultra or Centurion Max in winter oilseed rape.
•ALS herbicides-Pacifica Plus, Broadway Star or Monolith in winter wheat.
Glyphosate use before harvesting to kill established plants but this will not destroy the seeds which have set.Confirmation of resistant
Italian ryegrass
Italian ryegrass populations from eight fields were collected prior to harvest in 2020 as a part of the ECT nationwide grass weed survey. Four of these populations (R1 to R4) from Tipperary, Cork and Meath, which appeared to exhibit resistance, were subjected to detailed dose-response studies where they were compared with a known sensitive population.
Plants were sprayed at the two- to three-leaf stage, with rates ranging from 0.25 to eight times the recommended field rates of ALS (Pacifica Plus and Broadway Star) and ACCase (Axial Pro, Falcon and Stratos Ultra) herbicides to quantify resistance levels (Figure 1).
Figure 1: Symptoms of susceptible and resistant (R1 to R4) populations of Italian ryegrass 28-days post the application of ALS (A and B) and ACCase (C to E) herbicides at dose rates from 0.25 to 8-times the recommended field rates (which are highlighted in red). The extent of the green vegetation in each row is indicative of the difficulty in killing that population with differing rates of individual herbicides.
The results in Figure 1 show:
Figures 1 A and B show that Pacifica Plus and Broadway Star were virtually ineffective in all populations, confirming that R1 to R4 were ALS-resistant.Figure 1 C shows that R2 and R3 survived Axial application, while R1 and R4 were totally killed by Axial at the lowest dose rate used.Figure 1 D shows that three populations (R1, R3 and R4) were totally killed by Falcon at the recommended field rate, but R2 was not adequately controlled.Figure 1 E shows that all four populations (R1 to R4) were totally killed by Stratos Ultra used at the recommended field rate.
Figure 2: Pendimethalin-treated seedlings from a susceptible and two resistant populations (R1 and R4) had stunted shoots, indicating that they are highly pendimethalin-sensitive (ie no NTSR). In contrast, pendimethalin-treated plants from resistant populations R2 and R3 had developed normal shoots, indicating some degree of tolerance to pendimethalin and confirming potential NTSR.
In summary, populations R1 and R4 were ALS-resistant only. While populations R2 and R3 were ACCase- and ALS-resistant.
Nevertheless, ACCase-Stratos Ultra was highly effective on all four resistant populations. These findings demonstrate the difficulty in predicting cross-resistance, and stress the need to protect effective chemistry for critical use only.
Types of resistance evolving
in Irish Italian ryegrass
Following repeated application, Italian ryegrass develops herbicide resistance via two distinct mechanisms: (a) target-site resistance (TSR) where a plant changes the structure of its herbicide-binding site due to mutation(s) and this blocks herbicide activity or (b) non-target-site resistance (NTSR) where plants are able to degrade herbicides very rapidly before they reach the binding site.
To detect the involvement of TSR mutations, DNA was extracted from the resistant populations R1 to R4 using plant materials that survived ACCase/ALS herbicide application.
We found TSR as the key mechanism for ALS resistance in R1, R2 and R4. Even though R2 survived Axial or Falcon, no ACCase TSR detected. Similarly, R3 survived Pacifica Plus and Broadway Star, as well as Axial, but without ALS or ACCase TSR. These results speculate the possibility of NTSR in R2 and R3.To detect the involvement of NTSR or enhanced metabolism, we carried out a simple test using the pre-emergence herbicide pendimethalin, which is likely to affect the activity of other pre-emergence herbicides as well. However, resistance to pre-emergence herbicides used for grass weed control tends to be only partial and builds up relatively slowly.
Positive correlation between pendimethalin resistance and NTSR has been used for decades to screen UK grass weed populations.
A second test using a portable lateral flow device, which works on the same basis as a pregnancy test or COVID-19 antigen test, was used in parallel to detect the levels of herbicide-degrading enzymes GSTF1 (glutathione transferase phi (F class) 1) within 10 minutes (Figure 3).
Figure 3: Box plots of test line reading at 10 minutes from leaf protein extracted from Italian ryegrass plants. Each bar represents test line reading of five independent samples from each population. Different letter represents statistical significance (p<0.05). The test line reading of R3 was ˜2 times higher and statistically different to susceptible population, indicating NTSR development.
Higher levels of GSTF1 in the sample positively correspond to the existence of NTSR in the population.
Note: All NTSR work was performed at a leading UK institution, which has a unique molecular research capacity.
Using herbicide screening and molecular diagnostic tools, we identified and confirmed the development of NTSR alone or NTSR and TSR combinations in Irish Italian ryegrass populations (Table 1).
NTSR is a serious threat, as it affects multiple herbicides with different modes of action, leaving very limited chemical options for weed control.
High levels of control in these farms are needed to avoid further spread and buildup of resistance. All growers should address this weed using a zero-tolerance policy.
Resistance management and IWM
Remember, even though herbicides provide the most efficient and cost-effective means of managing weeds, resistance will make them obsolete if they are used as a sole weed control method without IWM.
Italian ryegrass is an increasing grass weed problem in Irish fields. It carries a very high risk of resistance development and some herbicide resistant strains have already been found in Irish fields.
Based on our knowledge of cross-resistance to key herbicides and the mechanisms involved, the IWM actions needed are as follows:
1 Farms where Italian ryegrass populations have confirmed or suspected herbicide resistance to ALS modes of action.
Pre-emergence herbicides in combination with cultural/non-chemical IWM like delayed autumn drilling, spring cropping, higher seed rates, ploughing, and hand-rogueing are critical to managing the soil seedbank population.No evidence of glyphosate resistance detected in R1 to R4. Therefore, glyphosate use on stubbles (at least 1.5 l/ha) may offer effective control but a follow-up treatment achieving high rates of control may be needed in the crop. For ALS-resistant populations (eg R1 and R4 in Table 1), adequate control may initially be achieved using ACCase (eg Axial Pro, Falcon and Stratos Ultra) herbicides.2 Farms where Italian ryegrass populations have confirmed or suspected herbicide resistance to ACCase and ALS modes of action.
Glyphosate use on stubbles (at least 1.5 l/ha) may offer effective control of young plants before planting.For ACCase/ALS-resistant populations (eg R2 and R3 in Table 1), ACCase-Stratos Ultra seems to be unaffected by NTSR and hence can be used to eliminate these multi-resistant types. However, the rotational restrictions around winter oilseed rape will limit the effectiveness of Stratos Ultra, hence the need to practice all and every cultural/non-chemical IWM to reduce the seed bank and prevent further seed return.General
prevention and weed management
In severely infested fields, extreme IWM measures such as cutting the crop for silage or spraying it off with glyphosate before seed return are important. Stale seedbed and cultivation: shallow cultivation immediately after harvest encourages germination of freshly shed seeds, which should be sprayed off with glyphosate prior to sowing. This technique is critical for direct drill or strip till, where seeds are retained on the soil surface. While ploughing can bury weed seeds to a depth from which they are unlikely to emerge, it can also bring previously buried seeds to the surface. Alternating the plough with a few years of minimum cultivation can reduce seedbank build-up.Delayed autumn drilling until mid-October will help to bypass the main autumn flush of Italian ryegrass and use higher seed rates to improve relative crop competitiveness.Spring cropping is a practical option where delayed autumn drilling is not possible or where weed pressure is significant
UK experience shows that hybrid barley can significantly reduce the level of Italian ryegrass seed return due to its higher competitive ability. Spring cropping is a practical option where delayed autumn drilling is not possible or where weed pressure is significant. However, a single spring crop may not be appropriate in fields with high soil seedbank populations. Including broadleaved crops in the rotation allows the use of other herbicide types (eg ACCase-Stratos Ultra). Hand-rogueing of small infestations or spraying off distinct patches prevent spreading within fields.Machine hygiene prevents weed seed movement within or between fields.Where infestations have occurred, pre-emergence herbicides should be followed by post-emergence herbicides applied to small actively growing plants. This practice should only be adopted in fields where weed pressure is low and ACCase/ALS resistance is not suspected.Finally, conduct resistance testing not only to determine whether the population is resistant, but also to identify the most effective chemistry that offers control for your populations.The presence of target site resistance and/or non-target site resistance has been confirmed in Irish Italian ryegrass populations.Increased vigilance, resistance testing and a zero-tolerance policy are critical to combat this weed.Do not use ACCase/ALS herbicides as a sole weed control method in consecutive crops due to its high resistance development risk.Use a wide range of cultural and non-chemical IWM options to protect effective herbicides for critical use and for the productivity of our cropping systems. 
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